C-reactive protein (CRP) is a prototype acute phase protein which increases dramatically in concentration in the blood during the first 24-48 hours of tissue necrosis and inflammation, Gewurz, Hospital Practice, 17, 67 (1982), and tests that measure CRP are used clinically to monitor the course of inflammatory reactions. CRP and/or CRP complexes can activate the complement system, Kaplan and Volanakis, J.Immunol, 112, 2135 (1974); Siegel et al., J. Exp. Medicine, 140, 631 (1974). They can also bind to and promote the activation of neutrophils and macrophages and enhance the respiratory burst response of these cells to certain stimuli in vitro. Mortensen et al., J. Immunol., 117, 774 (1976); Kilpatrick and Volanakis, J. Immunol., 134, 3364 (1985); Zeller et al., J. Lab. Clin. Med., 108, 567 (1986); Zeller et al., J. Leukocyte Biol., 40, 769 (1986). CRP has the capacity to protect against pneumococcal infections, influence clearance reactions, modify antibody formation to certain antigens and inhibit the metastasis of certain tumors in the mouse in vivo. Mold et al., J. Exp. Med., 154, 1703 (1981); Nakayama et al., J. Immunol. 128, 2435 (1982); Nakayama et al., J. Immunol , 54, 319 (1984); Deodhar et al., Cancer Res., 42, 5084 (1982). These properties suggest a significant biological role for CRP.
Native CRP is a cyclic pentamer composed of five identical, noncovalently-associated subunits, with its prototypic binding reactivity directed to phosphorylcholine (PC).sup.5 in the presence of calcium. Volanakis and Kaplan, Proc. Soc. Exp. Bio. Med. 136, 612 (1971). Recently in Potempa et al., Molec. Immunol., 20, 1165 (1983) and Potempa et al., Molec Immunol , 24, 531 (1987), a "neo-CRP" antigenicity was described. The neo-CRP antigenicity is expressed by native CRP modified by urea-chelation, acid treatment, heating, or direct immobilization on polystyrene plates The neo-CRP anti-genicty is also expressed on the intact CRP subunits and on the in vitro translation product of the CRP gene produced by hosts transformed with the gene by recombinant DNA techniques, rantzouranis et al., Pediatric Res. 18, 260a (1984).
The two molecular configurations of CRP can be distinguished antigenically, electrophoretically and by ligand binding reactivity. Potempa et al., Molec. Immunol., 20, 1165 (1983) and Potempa et al., Molec. Immunol., 24, 531 (1988). Polyclonal goat anti-native CRP reacted preferentially with both free and ligand-bound CRP in the presence of calcium (i.e., with native CRP epitopes), but showed no or minimal reactivity with CRP immobilized directly on solid phase surfaces or treated with urea, acid or heat in the absence of calcium. By contrast, polyclonal goat anti-neo-CRP showed preferential reactivity with CRP modified or immobilized in the absence of calcium (i.e., with neo-CRP epitopes) and little or no reactivity with CRP bound to ligands in the presence of calcium.
Forms of CRP expressing neo-CRP epitopes have the ability to activate and modulate the functional responses of platelets, polymorphonuclear leukocytes and monocytes, Potempa et al., Inflammation. 12, 391 (1988). Further, polyclonal antiserum to neo-CRP has been used to look for neo-CRP determinants in serum and tissue. Neo-CRP antigenicity has been found on the surface of human natural killer (NK) cells and B lymphocytes, Bray et al., J. Immunol , 140, 4271 (1988), as well as in human skeletal muscle tissue, Rees et al., Clin. Res., 37, 559a (1989). Potempa et al., Molec. Immunol., 24, 531 (1987) described neo-CRP reactivity in the serum or plasma of patients with rheumatoid arthritis. Rees et al., Clin. Immunol Immunopathol., 48, 95 (1988) demonstrated neo-CRP reactivities in frozen sections of acute phase but not normal rabbit liver and in necrotic but not normal rabbit muscle. The presence of neo-CRP determinants in acute phase liver was interpreted to result from de novo synthesis of the CRF subunit, since primary translation products of both human CRP (Mantzouranis et al., Pediatric Res., 18, 260a (1984)) and rabbit CRP (Samols et al., Biochem J., 227, 759 (1985)) had been shown to precipitate with a polyclonal antibody specific for neo-CRP epitopes. The expression of neo-CRP epitopes in necrotic tissue was attributed to in situ acute phase modification of the CRP molecule. These properties suggest a significant biological role for neo-CRP, as well as native CRP.
Additional articles describing neo-CRP and its properties include: Potempa et al, Protides of the Biological Fluids 34, 287 (1986); Bray et al., Clin. Immunol. Newsletter, 8, 137 (1987); Samberg et al., Cellular Immunology, in press; Gupta et al., Arthritis & Rheumatism 31, R39a (1988); Chu et al., Amer. Assoc. Cancer Res., 29, 371a (1988); Dougherty et al., Protides of the Biological Fluids, 34 291-93 (1986); and Chudwin et al., J. Allergy Clin. Immunol., 77, 2169 (1986).
Monoclonal antibodies to CRP prepared using native CRP as the immunogen are known. See Kilpatrick et al., Molec Immunol., 19, 1159 (1982); Tseng et al., Hybridoma, 7, 185 (1988); Kearney et al., Immunol Communications, 11, 275-82 (1982); Hirai et al., Protides of the Biological Fluids 34, 283 (1986); Tseng and Mortensen, Molecular Immunology, 25, 679 (1988); Roux et al., J. Immunol., 131 2411-15 (1983); Volanakis and Kearney, J. Exp. Med., 153, 1604 (1981). However, no reports describing monoclonal antibodies having the properties of the monoclonal antibodies of the present invention are known to Applicants. In particular, prior to the present invention, monoclonal antibodies reactive with modified CRP were not known.